Characterising the molecular response of Agaricus bisporus to mushroom virus X infection A thesis submitted to Maynooth University for the degree of Doctor of Philosophy Eoin O’Connor, B.Sc. October 2019 Supervisor Supervisor Head of Department Dr. David Fitzpatrick Dr. Helen Grogan Prof. Paul Moynagh Department of Biology Horticulture Development Department of Biology Department Maynooth University Maynooth University Teagasc Food Research Co. Kildare Co. Kildare Centre Ireland Ireland Ashtown, Dublin 8 Ireland Table of Contents Index of Figures ................................................................................................................................... III Index of Tables ..................................................................................................................................... V Abbreviations ....................................................................................................................................... VI Acknowledgements .............................................................................................................................. X Declaration .......................................................................................................................................... XII Statement of Contributions ............................................................................................................... XII Publications and Presentations ........................................................................................................ XIII Publications ....................................................................................................................................... XIII Abstract ............................................................................................................................................... XVI Preface ............................................................................................................................................... XVII 1.0 ....................................................................................................................... General Introduction ................................................................................................................................................................ 2 1.1 A history of Agaricus bisporus .................................................................................................... 2 1.2 Biology of A. bisporus .................................................................................................................. 4 1.3 Commercial Mushroom Breeding ............................................................................................... 9 1.4 Mushroom Browning .................................................................................................................. 11 1.5 Cultivation of Agaricus bisporus ............................................................................................... 13 1.6 Pests and pathogens of A. bisporus ........................................................................................... 18 1.7 Mycoviruses ................................................................................................................................. 21 1.8 Mycoviruses of A. bisporus ........................................................................................................ 26 1.9 Host mycoviral defence .............................................................................................................. 34 1.10 Functional analyses of the A. bisporus genome .................................................................... 36 1.11 Aims and Objectives ................................................................................................................. 40 2.0 .......... Whole Genome Sequence of the Commercially Relevant Mushroom Strain Agaricus bisporus var. bisporus ARP23 .......................................................................................................... 57 Abstract ................................................................................................................................................ 58 2.1 Introduction .................................................................................................................................. 59 2.2 Methods ......................................................................................................................................... 61 2.3 Results and Discussion ................................................................................................................ 67 2.4 Conclusion .................................................................................................................................... 79 Literature Cited ................................................................................................................................... 80 3.0 ... Proteomic investigation of interhyphal interactions between strains of Agaricus bisporus .............................................................................................................................................................. 88 Abstract ................................................................................................................................................ 89 I 3.1 Introduction .................................................................................................................................. 90 3.2 Methods ......................................................................................................................................... 94 3. Results ............................................................................................................................................. 97 3.4 Discussion ................................................................................................................................... 117 3.5 Conclusion .................................................................................................................................. 121 Literature Cited ................................................................................................................................. 122 4.0 ........................ Transmission of Mushroom Virus X and the impact of virus infection on the transcriptomes and proteomes of different strains of Agaricus bisporus ................................. 134 Abstract .............................................................................................................................................. 135 4.1 Introduction ................................................................................................................................ 136 4.2 Methods ....................................................................................................................................... 139 4.3. Results ........................................................................................................................................ 145 4.4 Discussion ................................................................................................................................... 188 5. Conclusion .................................................................................................................................... 196 Literature Cited ................................................................................................................................. 197 5.0 ............. FISHing in fungi: Visualisation of mushroom virus X in the mycelium of Agaricus bisporus by fluorescence in situ hybridisation ............................................................................. 204 Abstract .............................................................................................................................................. 205 5.1 Introduction ................................................................................................................................ 206 5.2 Methods ....................................................................................................................................... 208 5.3 Results ......................................................................................................................................... 213 5.4 Discussion ................................................................................................................................... 219 5. Conclusion .................................................................................................................................... 221 Literature Cited ................................................................................................................................. 222 6.0 ................................................................................................... Synthesis and Future Directions ............................................................................................................................................................ 226 Literature Cited ................................................................................................................................. 238 Supplementary Information ............................................................................................................ 241 II Index of Figures Chapter 1 Figure 1.1: The life cycle of Agaricus bisporus var. bisporus. 8 Figure 1.2: